Sudden arrhythmia death syndrome in young victims: a five-year retrospective review and two-year prospective molecular autopsy study by next-generation sequencing and clinical evaluation of their first-degree relatives.

OBJECTIVE: Sudden arrhythmia death syndrome (SADS) accounts for about 30% of causes of sudden cardiac death (SCD) in young people. In Hong Kong, there are scarce data on SADS and a lack of experience in molecular autopsy. We aimed to investigate the value of molecular autopsy techniques for detecting SADS in an East Asian population. METHODS: This was a two-part study. First, we conducted a retrospective 5-year review of autopsies performed in public mortuaries on young SCD victims. Second, we conducted a prospective 2-year study combining conventional autopsy investigations, molecular autopsy, and cardiac evaluation of the first-degree relatives of SCD victims. A panel of 35 genes implicated in SADS was analysed by next-generation sequencing. RESULTS: There were 289 SCD victims included in the 5-year review. Coronary artery disease was the major cause of death (35%); 40% were structural heart diseases and 25% were unexplained. These unexplained cases could include SADS-related conditions. In the 2-year prospective study, 21 SCD victims were examined: 10% had arrhythmogenic right ventricular cardiomyopathy, 5% had hypertrophic cardiomyopathy, and 85% had negative autopsy. Genetic analysis showed 29% with positive heterozygous genetic variants; six variants were novel. One third of victims had history of syncope, and 14% had family history of SCD. More than half of the 11 first-degree relatives who underwent genetic testing carried related genetic variants, and 10% had SADS-related clinical features. CONCLUSION: This pilot feasibility study shows the value of incorporating cardiac evaluation of surviving relatives and next-generation sequencing molecular autopsy into conventional forensic investigations in diagnosing young SCD victims in East Asian populations. The interpretation of genetic variants in the context of SCD is complicated and we recommend its analysis and reporting by qualified pathologists.

Initial experience of cryoballoon catheter ablation for atrial fibrillation in Hong Kong.

OBJECTIVE. To report the initial experience in using cryoballoon catheter ablation in the treatment of atrial fibrillation in Hong Kong. DESIGN. Single-centre, prospective case series. SETTING. Regional hospital, Hong Kong. PATIENTS. Sixteen patients (mean age, 55 years; standard deviation, 14 years; 11 males) with paroxysmal (n=12) or persistent (n=4) atrial fibrillation. INTERVENTIONS. Pulmonary vein isolation by ablation with a 28-mm cryoballoon catheter. MAIN OUTCOME MEASURES. Safety, effectiveness, and learning curve of this procedure. RESULTS. Of 67 pulmonary veins, 61 (91%) could be successfully isolated with the cryoballoon alone. The remaining pulmonary veins were isolated with additional ablation using an 8-mm tip cryocatheter. One phrenic nerve palsy developed during right middle pulmonary vein ablation, which resolved. Another patient endured a minor guidewire dissection of the right inferior pulmonary vein. The mean (standard deviation) procedural and fluoroscopic times were 231 (32) and 62 (18) minutes, respectively. On comparing the first nine and last seven procedures, there was a significant improvement in procedural time (mean [standard deviation], 244 [32] vs 213 [24] minutes; P=0.04) and in the fluoroscopic time (70 [21] vs 51 [7] minutes; P=0.038). With a median follow-up of 21 months, nine (75%) of the 12 patients with paroxysmal atrial fibrillation and one (25%) of those four with persistent atrial fibrillation had no recurrence, without the use of anti-arrhythmic drugs. CONCLUSIONS. Pulmonary vein isolation by cryoballoon catheter ablation is safe and effective in treating patients with paroxysmal, but not for patients with persistent atrial fibrillation. A relatively short learning curve of around 10 cases was deemed appropriate.

Ceramide inhibits the inwardly rectifying potassium current in GH(3) lactotrophs.

The effects of ceramide on ion currents in rat pituitary GH(3) cells were investigated. Hyperpolarization-elicited K(+) currents present in GH(3) cells were studied to determine the effect of ceramide and other related compounds on the inwardly rectifying K(+) current (I(K(IR))). Ceramide (C(2)-ceramide) suppressed the amplitude of I(K(IR)) in a concentration-dependent manner, with an IC(50) value of 5 microM. Ceramide caused a rightward shift in the midpoint for the activation curve of I(K(IR)). Pretreatment with PD-98059 (30 microM) or U-0126 (30 microM) did not prevent ceramide-mediated inhibition of I(K(IR)). However, the magnitude of ceramide-induced inhibition of I(K(IR)) was attenuated in GH(3) cells preincubated with dithiothreitol (10 microM). TNF alpha (100 ng/g) also suppressed I(K(IR)). In the inside-out configuration, application of ceramide (30 microM) to the bath slightly suppressed the activity of large conductance Ca(2+)-activated K(+) channels. Under the current clamp mode, ceramide (10 microM) increased the firing of action potentials. Cells that exhibited an irregular firing pattern were converted to those displaying a regular firing pattern after application of ceramide (10 microM). Ceramide also suppressed I(K(IR)) in neuroblastoma IMR-32 cells. Therefore, ceramide can produce a depressant effect on I(K(IR)). The blockade of this current by ceramide may affect cell function.

Postanoxic parkinsonism: clinical, radiologic, and pathologic correlation.

The authors report a 72-year-old patient who presented with parkinsonism after hypoxic-ischemic insult. T1-weighted MRI revealed high signal intensity lesions in the basal ganglia. Pathologic study of the brain disclosed multiple foci of old infarcts with gliosis and lipid-laden and hemosiderin-laden macrophages, indicating a previous minor hemorrhage after infarction. This observation provided pathologic correlation with the patient's clinical symptoms and MRI.

Oral drug of choice in carpal tunnel syndrome.

BACKGROUND: Conservative treatment of mild to moderate carpal tunnel syndrome (CTS) is variable. OBJECTIVE: To evaluate the effectiveness of commonly used oral medications such as diuretics, nonsteroid anti-inflammatory drugs (NSAIDs), and steroids in the treatment of CTS. METHODS: Prospective, randomized, double-blind and placebo-controlled study of patients with clinical symptoms and signs of CTS, confirmed by standard electrodiagnosis. Baseline assessments included a standardized symptom questionnaire, rating five categories of symptoms (pain, numbness, paresthesia, weakness/clumsiness, and nocturnal awakening) on a scale from 0 (no symptoms) to 10 (severe). The total score in each of the five categories was termed the global symptom score (GSS). After baseline assessment, patients were randomized to the following treatment arms: 1) 4 weeks of placebo (n = 16); 2) 4 weeks of diuretic (trichlormethiazide, 2 mg daily; n = 16); 3) 4 weeks of NSAID-slow release (SR) (tenoxicam-SR, 20 mg daily; n = 18); and 4) 2 weeks of prednisolone, 20 mg daily, followed by another 2-week dosage of 10 mg daily (n = 23). Results of follow-up assessments in the second and the fourth weeks were identical to baseline scores. The changes in GSS were analyzed to determine the statistical difference. RESULTS: No significant reduction from baseline GSS was seen at second, and fourth weeks in the placebo, NSAID-SR, and diuretic groups. However, the mean score at 4 weeks in the steroid group decreased significantly from a baseline of 27.9 +/- 6.9 to 10 +/- 7.4. CONCLUSION: For patients with mild to moderate CTS who opt for conservative treatment, corticosteroids are of greater benefit.

Hypertrophic cranial pachymeningitis and lymphocytic hypophysitis in Sjögren's syndrome.

The authors describe a patient with primary Sjögren's syndrome who developed pachymeningitis, hypopituitarism, and central diabetes insipidus. The patient improved with corticosteroid pulse therapy.

Rutaecarpine-induced block of delayed rectifier K+ current in NG108-15 neuronal cells.

The effects of rutaecarpine on ionic currents of NG108-15 neuronal cells were investigated in this study. Rutaecarpine (2-100 microM) suppressed the amplitude of delayed rectifier K+ current (I(K(DR))) in a concentration-dependent manner. The IC50 value for rutaecarpine-induced inhibition of I(K(DR)) was 11 microM. I(K(DR)) present in these cells is sensitive to the inhibition by quinidine and dendrotoxin, yet not by E-4031. The presence of rutaecarpine enhanced the rate and extent of I(K(DR)) inactivation, although it had no effect on the initial activation phase of I(K(DR)). Recovery from block by rutaecarpine (5 microM) was fitted by a single exponential with a value of 2.87 s. Crossover of tail currents in the presence of rutaecarpine was also observed. Cell-attached single-channel recordings revealed that rutaecarpine decreased channel activity, but it did not alter single-channel amplitude. With the aid of the binding scheme, a quantitative description of the rutaecarpine actions on I(K(DR)) was provided. However, rutaecarpine (20 microM) had no effect on L-type Ca2+ current. Under current-clamp configuration, rutaecarpine prolonged action potential duration in NG108-15 cells. These results show that rutaecarpine is a blocker of the K(DR) channel. The increase in action potential duration induced by rutaecarpine can be explained mainly by its blocking actions on I(K(DR)).

Mechanisms of histamine-induced intracellular Ca2+ release and extracellular Ca2+ entry in MG63 human osteosarcoma cells.

The effect of histamine on intracellular free Ca2+ levels ([Ca2+](i)) in MG63 human osteosarcoma cells was explored using fura-2 as a Ca2+ dye. Histamine increased ([Ca2+](i)) in a concentration-dependent fashion with an EC(50) value of 0.5 microM. Extracellular Ca2+ removal inhibited the ([Ca2+](i)) signals. Histamine failed to increase ([Ca2+](i)) in Ca2+-free medium after cells were pretreated with thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor). Addition of Ca2+ induced concentration-dependent ([Ca2+](i)) increases after preincubation with histamine in Ca2+-free medium. Histamine-induced intracellular Ca2+ release was abolished by inhibiting phospholipase C with 1-(6-((17beta-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione (U73122). The ([Ca2+](i)) increase induced by histamine in Ca2+ medium was abolished by cimetidine, but was not altered by pyrilamine, nifedipine, verapamil, and La(3+). Together, this study shows that histamine increased in ([Ca2+](i)) in osteosarcoma cells by stimulating H2 histamine receptors. The Ca2+ signal was caused by Ca2+ release from the endoplasmic reticulum in a phospholipase C-dependent manner. The Ca2+ release was accompanied by Ca(2+) influx.

Effect of oleamide on Ca(2+) signaling in human bladder cancer cells.

The effect of oleamide, a sleep-inducing endogenous lipid in animal models, on intracellular free levels of Ca(2+) ([Ca(2+)](i)) in non-excitable and excitable cells was examined by using fura-2 as a fluorescent dye. [Ca(2+)](i) in pheochromocytoma cells, renal tubular cells, osteoblast-like cells, and bladder cancer cells were increased on stimulation of 50 microM oleamide. The response in human bladder cancer cells (T24) was the greatest and was further explored. Oleamide (10-100 microM) increased [Ca(2+)](i) in a concentration-dependent fashion with an EC(50) of 50 microM. The [Ca(2+)](i) signal comprised an initial rise and a sustained plateau and was reduced by removing extracellular Ca(2+) by 85 +/- 5%. After pre-treatment with 10-100 microM oleamide in Ca(2+)-free medium, addition of 3 mM Ca(2+) increased [Ca(2+)](i) in a manner dependent on the concentration of oleamide. The [Ca(2+)](i) increase induced by 50 microM oleamide was reduced by 100 microM La(3+) by 40%, but was not altered by 10 microM nifedipine, 10 microM verapamil, and 50 microM Ni(2+). In Ca(2+)-free medium, pre-treatment with thapsigargin (1 microM), an endoplasmic reticulum Ca(2+) pump inhibitor, abolished 50 microM oleamide-induced [Ca(2+)](i) increases; conversely, pretreatment with 50 microM oleamide reduced 1 microM thapsigargin-induced [Ca(2+)](i) increases by 50 +/- 3%. Suppression of the activity of phospholipase C with 2 microM U73122 failed to alter 50 microM oleamide-induced Ca(2+) release. Linoleamide (10-100 microM), another sleep-inducing lipid with a structure similar to that of oleamide, also induced an increase in [Ca(2+)](i). Together, it was shown that oleamide induced significant [Ca(2+)](i) increases in cells by a phospholipase C-independent release of Ca(2+) from thapsigargin-sensitive stores and by inducing Ca(2+) entry.

Fendiline, an anti-anginal drug, increases intracellular Ca2+ in PC3 human prostate cancer cells.

BACKGROUND: The effects of the anti-anginal drug fendiline on intracellular Ca2+ concentrations ([Ca2+]i) in human PC3 prostate cancer cells were examined. METHODS: [Ca2+]i was measured using the fluorescent dye fura-2. RESULTS: Fendiline (0.5-100 microM) increased [Ca2+]i in a concentration-dependent manner. Ca2+ removal partly inhibited the Ca2+ signals. In Ca2+-free medium, pretreatment with 100 microM fendiline inhibited most of the [Ca2+]i increase induced by 1 microM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor), and pretreatment with thapsigargin abolished the fendiline-induced [Ca2+]i increases. Adding 3 mM Ca2+ increased [Ca2+]i in cells pretreated with 0.5-200 microM fendiline in Ca2+-free medium. Pretreatment with 1 microM U73122 to block the formation of inositol-1.4.5-trisphosphate (IP3) did not alter fendiline-induced internal Ca2+ release. CONCLUSIONS: The anti-anginal drug fendiline induced internal Ca2+ release and external Ca2+ entry. Because prolonged increases in [Ca2+]i may lead to cell injury and death, the long-term effect of fendiline on the function of prostate cancer cells should be investigated.

The cause of slowed forearm median conduction velocity in carpal tunnel syndrome.

OBJECTIVES: Attempting to answer a debate concerning the etiopathogenesis of the decreased forearm median motor conduction velocity (FMMCV), we tried to use proximal stimulation at the wrist, elbow, mid-arm and axillary regions to determine segmental median motor conduction velocity (MMCV). We also correlated the FMMCV with median motor distal latency (MMDL) and compound muscle action potential (CMAP) amplitudes of the abductor pollicis brevis (APB) muscle in order to assess whether the conduction block of large myelinating fibers or retrograde axonal atrophy was the major cause of the decreased FMMCV. BACKGROUND: The cause of the decreased FMMCV resulting from either the conduction block of the large myelinating fibers at the wrist or distal compression with retrograde axonal atrophy remains an unresolved issue at the moment. Animal models have supported the hypothesis that the retrograde axonal atrophy might also occur in humans. Other authors believe the standard FMMCV is calculated by subtracting the distal latency which may not represent an exact assessment of FMMCV but rather the velocity of small fibers that persist through the carpal tunnel. SUBJECTS AND METHODS: Patients with the clinical symptoms and signs of carpal tunnel syndrome (CTS) confirmed using standard electrodiagnosis were included. The patients were arbitrarily divided into two groups based on the FMMCV, one with reduced FMMCV (n = 20, FMMCV < 50 m/s) and the other with normal FMMCV (n = 35, FMMCV> or =50 m/s). Age-matched volunteers served as controls. We explored motor conduction proximally at wrist, elbow, mid-arm and axillary stimulation, and recorded at the APB muscles. Based on the latency differences, we calculated the FMMCV, distal arm MMCV (DAMMCV) and proximal arm MMCV (PAMMCV), and compared the conduction velocity (CV) differences of DAMMCV-FMMCV, PAMMCV-FMMCV and PAMMCV-DAMMCV in the two patient groups and the control. Furthermore, we correlated FMMCV with MMDL and CMAP amplitudes of APB muscle because MMDL and CMAP amplitudes might reflect the integrity of the large myelinating fibers. RESULTS: CMAP amplitudes of APB muscle at wrist stimulation and MMDL were not correlated with FMMCV in either of the two patient groups; however, the CMAP amplitude was markedly decreased and MMDL was significantly prolonged when compared with normal controls. The significant increase of CV gradient of DAMMCV-FMMCV and PAMMCV-FMMCV without an equal increase of CV gradient of PAMMCV-DAMMCV only occurred in the reduced FMMCV patient group, suggesting that the conduction block is not the primary cause. The CV gradient of DAMMCV-FMMCV and PAMMCV-DAMMCV did not show any significant difference between patients with the normal FMMCV and the control group. CONCLUSION: The retrograde axonal atrophy, not selective damage of the large fibers at the wrist, was the direct cause of the decreased FMMCV.

Gossypol, a component in cottonseed, induced increases in cytosolic Ca2+ levels in Chang liver cells.

The effect of gossypol, a compound found in cottonseed, on intracellular free Ca2+ levels ([Ca2+](i)) in Chang liver cells were evaluated using fura-2 as a fluorescent Ca2+ indicator. Gossypol (0.2-5microM) increased [Ca2+](i) in a concentration-dependent manner with an EC(50) value of 1.5microM. The [Ca2+](i) response was composed of an initial rise and a slow decay to a sustained phase within 5min after drug application. Removal of extracellular Ca2+ markedly reduced the [Ca2+](i) signals by 80+/-2%. Preincubation with 0.1mM La3+ or 10microM nimodipine abolished the Ca2+ influx. Gossypol (5microM)-induced release of intracellular Ca2+ was reduced by 75% by pretreatment with 1microM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor) to deplete the endoplasmic reticulum Ca2+. Conversely, pretreatment with gossypol abolished thapsigargin-induced Ca2+ release. After pretreatment with 5microM gossypol in Ca2+-free medium for several min, addition of 3mM Ca2+ induced a [Ca2+](i) increase of a magnitude nine-fold greater than control. Gossypol (5microM)-induced Ca2+ release was not affected by inhibiting phospholipase C with 2microM 1-(6-((17beta-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione (U73122). Together, this study shows that gossypol induced significant [Ca2+](i) increases in Chang liver cells by releasing Ca2+ from intracellular pools in a phospholipase C-dissociated fashion and by causing La3+- and nimodipine-sensitive Ca2+ influx.

Spinal cord proliferative sparganosis in Taiwan: a case report.

A 43-year-old woman suffered from low back pain and bilateral footdrop. A cisternal myelogram unexpected revealed multiple filing defects in the spinal canal extending from the lower cervical region to the caudal equina. Diagnostic exploration revealed numerous cystic organisms adhering to the spinal cord and nerve roots. Histopathological examination showed these organisms to be proliferative sparganum cestode larvae. Although these cestode larval infections have been reported a dozen times in humans from various parts of the world, this is probably the first reported case of spinal cord infection.

Brain SPECT imaging with 99Tcm-hexamethylpropyleneamine oxime in the early detection of cerebral infarction: comparison with transmission computed tomography.

A new cerebral blood flow agent, 99Tcm-hexamethylpropyleneamine oxime (HM-PAO), was evaluated for early detection of acute cerebral infarction in conjunction with the transmission computed tomographic (CT) studies. The data from 22 cases were analysed. Results reveal that 99Tcm-HM-PAO enables the early detection of acute cerebral infarction prior to CT with rather proper depiction of the extent of physiological abnormality in the majority of patients. This promising result together with the lack of logistical problems will make 99Tcm-HM-PAO a useful and practical agent worldwide for diagnosing and managing acute cerebral infarction.

Effects of the antianginal drug fendiline on Ca2+ movement in hepatoma cells.

This study investigated the effect of the anti-anginal drug, fendiline, on intracellular free Ca2+ levels ([Ca2+]i) in HA/ 22 human hepatoma cells by using fura-2 as a fluorescent Ca2+ dye. Fendiline (1-100 microM) increased [Ca2+]i with an EC50 of 25 microM. Removal of extracellular Ca2+ reduced the [Ca2+]i signals by 51 +/- 5%. Fendiline (10 microM)-induced Ca2+ release was abolished by pretreatment with 1 microM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor). Inhibition of phospholipase C with 2 microM 1-(6-((17beta-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione (U73122) did not alter 10 microM fendiline-induced Ca2+ release. Several other calmodulin antagonists, such as phenoxybenzamine (100-200 microM), trifluoperazine (5-50 microM), and fluphenazine-N-chloroethane (2-100 microM), had no effect on [Ca2+]i. Together, it was found that fendiline increased [Ca2+]i in human hepatoma cells by discharging Ca2+ from the endoplasmic reticulum in an inositol 1,4,5-trisphosphate-independent manner and by inducing Ca2+ entry. This effect of fendiline does not appear to be via antagonism of calmodulin.

Sensitivity of diffusion-weighted magnetic resonance imaging in the diagnosis of acute lacunar infarcts.

BACKGROUND AND PURPOSE: Heightened interest in the early diagnosis and treatment of acute stroke challenges neuroimaging specialists to optimize available modalities and to develop new techniques for the evaluation of cerebrovascular disease. The purpose of this study was to evaluate the sensitivity of diffusion-weighted (DW) magnetic resonance (MR) imaging in detecting early small infarcts and in differentiating acute from nonacute small infarcts when conventional MR imaging demonstrates multiple small infarcts. METHODS: Thirty-eight consecutive patients with a clinical diagnosis of lacunar infarcts (20 men and 18 women, aged 50-79 yr) who underwent DW MR imaging within 3 days of symptom onset were enrolled in this study. All patients underwent both conventional fast spin-echo (FSE) MR imaging and DW MR imaging. Apparent diffusion coefficient (ADC) maps were also acquired. All patients had at least one of the following classic lacunar syndromes: pure motor hemiparesis, ataxic hemiparesis, dysarthria-clumsy hand, pure sensory stroke, and sensorimotor stroke. RESULTS: Thirty-six patients (40 acute lesions) had focal areas of high intensity on DW MR imaging associated with their clinical symptoms. Acute lacunar infarcts were seen on DW MR imaging as bright areas of decreased ADC ratio (range 0.31-0.85, mean 0.64). Lesion conspicuity with DW MR imaging was superior to that with FSE in 33 acute lesions. In four patients with small hyperacute (within 6 hours) infarcts, DW MR imaging was particularly sensitive for infarcts that were not visible on FSE sequences. The sensitivity of DW MR imaging and ADC map for acute lacunar infarcts was 95%, specificity 94%, positive predictive value 97%, negative predictive value 90%, and accuracy 95%. In 15 patients with both acute and nonacute old small infarcts, DW MR imaging and ADC map could easily distinguish the new infarct from adjacent old ones, although this distinction was difficult to make with FSE. CONCLUSIONS: DW MR imaging accompanied by ADC map is a sensitive diagnostic modality for hyperacute and acute lacunar infarcts. It is also sensitive in distinguishing fresh small infarcts from adjacent multiple old infarcts.

Functional coupling of voltage-dependent L-type Ca2+ current to Ca2+-activated K+ current in pituitary GH3 cells.

Ca2+-activated K+ currents (I(K(Ca)) can contribute to action potential repolarization and after-hyperpolarization in GH3 cells. In this study, we examined how the activation of I(K(Ca) at the cellular level could be functionally coupled to Ca2+ influx through L-type Ca2+ channels. A 30-msec Ca2+ influx step to 0 mV was found to exhibit substantial contribution of Ca2+ influx through the activation of I(Ca,L) to the activation of I(K(Ca)). A bell-shaped relationship between the conditioning potentials and the integrated I(K(Ca)) was observed, suggesting that the magnitude of integrated I(Ca,L) correlates well with that of integrated I(K(Ca)) in the same cell. A linear relationship of integrated I(Ca,L) and integrated I(K(Ca)) was found with a coupling ratio of 69+/-7. The value of the coupling ratio was unaffected by the presence of Bay K 8644 or nimodipine, although these compounds could effectively affect the amplitudes of both I(K(Ca)) and I(Ca,L). However, tetrandrine could decrease the coupling ratio. Paxilline or intracellular Ca2+ buffer with EGTA decreased the coupling ratio, while apamin had no effect on it. Interestingly, phorbol 12-myristate 13-acetate also reduced the coupling ratio significantly, whereas thapsigargin increased this value. Thus, the present study indicates that the activation of I(K(Ca)) during brief Ca2+ influx, which is inhibited by paxilline, is coupled to Ca2+ influx primarily through the L-type channels. The selective modulation of I(K(Ca)) by second messengers or Ca2+ release from internal stores may affect the coupling efficiency and hence cellular excitability.

Fendiline-Induced Ca2+ movement in A10 smooth muscle cells.

The effect of fendiline, an anti-anginal drug, on cytosolic free Ca2+ levels ([Ca2+]i) in A10 smooth muscle cells was explored by using fura-2 as a Ca2+ indicator. Fendiline at concentrations between 10-50 microM increased [Ca2+]i in a concentration-dependent manner with an EC50 of 20 microM. External Ca2+ removal reduced the Ca2+ signal by 75%. Addition of 3 mM Ca2+ increased [Ca2+]i in cells pretreated with fendiline in Ca2+-free medium. The 50 microM fendiline-induced [Ca2+]i increase in Ca2+-containing medium was inhibited by 10 microM of La3+, nifedipine, or verapamil. In Ca2+-free medium, pretreatment with 1 microM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor) to deplete the endoplasmic reticulum Ca2+ store partly inhibited 50 microM fendiline-induced Ca2+ release; whereas pretreatment with 50 microM fendiline abolished 1 microM thapsigargin-induced Ca2+ release. Inhibition of phospholipase C activity with 2 microM U73122 did not alter 50 microM fendiline-induced Ca2+ release. Incubation with 50 microM fendiline for 10-30 min decreased cell viability by 10-20%. Together, the findings indicate that in smooth muscle cells fendiline induced [Ca2+]i increases. Fendiline acted by activating Ca2+ influx via L-type Ca2+ channels, and by releasing internal Ca2+ in a phospholipase C-independent manner. Prolonged exposure of cells to fendiline induced cell death.

Mechanism of bifonazole-induced [Ca2+]i increases in MDCK renal tubular cells.

The effect of the antifungal drug bifonazole on Ca2+ homeostasis in Madin Darby canine kidney (MDCK) cells was investigated. Cell suspensions were loaded with the Ca2+-sensitive dye fura-2, and the fluorescence changes were measured with a spectrofluorophotometer. At concentrations between 10-80 microM bifonazole increased cytosolic free Ca2+ levels ([Ca2+]i) in a concentration-dependent manner. The Ca2+ signals were partly inhibited by removing extracellular Ca2+. Bifonazole (40 microM) released Ca2+ from the store sensitive to 1 microM thapsigargin, an endopolasmic reticulum Ca2+ pump inhibitor. Bifonazole (40 microM) per se induced capacitative Ca2+ entry while reduced 1 microM thapsigargin-induced capacitative Ca2+ entry. Inositol 1,4,5-trisphosphate may be involved in bifonazole-induced Ca2+ release because inhibiting phospholipase C with 2 microM U73122 partly reduced the bifonazole response. Together, bifonazole increased [Ca2+]i in renal tubular cells by inducing intracellular Ca2+ release and extracellular Ca2+ influx.